Within each cell type, organelles and macromolecular complexes are localized to specific locations. The movements of these subcellular components are regulated both temporally and spatially and generally occur via microtubule and actin-based movement. One essential form of actin-based movement is dependent on Class V myosins. The overall goal of this study is to determine how class V myosins attach and detach from cargoes at the correct time and place. This question is of critical interest because most myosin V motors, including a Saccharomyces cerevisiae myosin V, Myo2, move multiple cargoes. Each type of cargo has its own pattern of cell-cycle coordinated movement. Furthermore, the ultimate destinations of each cargo are not the same. Insights into the regulation of myosin V attachment and detachment have come from our discovery that Vacl? and Vac8 are key components of a vacuole (lysosome)-specific myosin receptor. Initiation of vacuole inheritance requires Vacl7/Myo2 interactions. In addition, when vacuoles reach their proper destination, Vacl 7 is specifically degraded, depositing the vacuoles in their correct location. Our major goals are to 1) determine the molecular basis of Vac8, Vacl7 and Myo2 interactions, 2) determine how the regulated turnover of Vac 17 occurs and whether the signal for Vac 17 turnover is spatial or temporal, 3) determine how the initiation and termination of vacuole inheritance are coordinated with the cell-cycle, and 4) identify other members of the transport complex. These studies should reveal how regulation of both myosin V and its receptor move cargo to the correct place at the proper time. This question is of clear relevance to human health because the homologue of yeast Myo2, myosin Va, is required for correct positioning of synaptic vesicles, the endoplasmic reticulum, and melanosomes. Myosin Va is essential for human life. Patients with partially functional myosin Va, have Griscelli's syndrome, and display both neurological and pigmentation defects. [unreadable] [unreadable]